Respiratory tract infections are a leading cause of premature mortality in the US and worldwide. Even upper respiratory infections, such as otitis media and sinusitis can lead to morbidity. Consequently, innate immunity is particularly important component of host defense in the human respiratory tract, particularly in the upper airway which is heavily colonized with a variety of commensal microorganisms, as well as intermittent colonization with known respiratory tract pathogens such as Streptococcus pneumoniae and Haemophilus influenzae that commonly coexist on the mucosal surface of the nasopharynx. Mediators of host immunity in the human respiratory tract include immunoglobulins A and G, lysozyme, lactoferrin, mucus glycoproteins, secretory leukoprotease inhibitor, secretory phospholipase A, uric acid, peroxidase, aminopeptidase and neutral endopeptidase (Kaliner, Am. Rev. Respir. Dis. 144:S52 (1991); Kim & Weiser, “Medical Importance of Normal Microflora,” in Respiratory Infections, (Tannock, ed.) Thompson Science, London, 1998). As a result, it is often difficult to for healthcare personnel to distinguish between those microorganisms that are causing infection, from those that are simply colonizing the region.
C-reactive protein (CRP) is a known constituent of human sera (Szalai et al., Immunol. Res. 16:127-136 (1997)), produced by hepatocytes as a single chain precursor with a cleavable signal sequence at the N-terminus (Weiser et al., J. Exp. Med. 187:631-640 (1998)), and induced by proinflammatory cytokines. CRP derives its name from the finding that it binds to the C polysaccharide or cell wall teichoic acid of Streptococcus pneumoniae. The concentration of CRP in serum is generally less than 2 μg/ml, but it is known to increase to as much as 1000-fold in response to a stimulus, such as tissue injury or inflammation to levels typically >50 μg/ml (Claus et al., J. Lab. Clin. Med. 87:120-128 (1976)). Moreover, the CRP levels are known to decline rapidly when the inflammatory stimulus has been removed with a half-life of ˜4-7 hours (Pepys et al., Adv. Immunol. 34:141 (1983); Young et al., Pathology 23:118 (1991)). As a result, CRP is a recognized clinical marker of the inflammatory process in vivo, although the function of this acute-phase reactant and its precise role in host defense remain poorly understood (Pepys et al., Nature 278:259-261 (1979); Whitehead et al., Biochem. J. 266:283-290 (1990)).
Mice, which have a constitutively low level of CRP, are resistant to experimental pneumococcal sepsis when carrying the human CRP transgene at levels that would otherwise confer inducible high-levels of expression (Szalai et al., J. Immunol. 155:2557-2563 (1996)). Therefore, the use of animal models has been limited because the regulation of CRP in such animals simply does not parallel that of humans.
Human serum CRP is a cyclic pentameric protein (˜120 kD), having five, identical, non-covalently bound, non-glycosylated subunits of 206 amino acids, each having a molecular mass of 24 kDa. (Gerwurz et al., Curr. Opin. Immunol. 7:54-64 (1995)). CRP binds in a calcium-dependent manner to choline phosphate or phosphorylcholine (ChoP) residues found on C polysaccharide (Volanakis et al., Proc. Soc. Exp. Biol. Med. 136:612 (1971)). ChoP is considered to be a highly usual structural feature in prokaryotes. However, in addition to S. pneumoniae, many of the bacteria that normally inhabit the human respiratory tract express ChoP, the molecular target of CRP, on their cell surface (Mosser et al., J. Biol. Chem. 245:287-298 ((1970)). Found primarily on the mucosal surface of the airway, ChoP has been found on the cell surface of both gram-positive and gram-negative species of bacteria, and it has also been found on Mollicutes. More specifically, it is known to be present on the cell surface of certain bacterial pathogens of the human respiratory tract, including Streptococcus oralis, S. mitis, Haemophilus influenzae, H. somnus, Actinobacillus actinomycetemcomitans, Pseudomonas aeruginosa, and Fusobacterium nucleatum, as well as certain species of Actinomyces, Neisseria, such as N. meningitides, and Mycoplasma, such as M. fermentans and M. pneumoniae. 
However, in an analysis of respiratory tract surface exudates, CRP was not recognized as a component of innate antimicrobial activity (Cole et al., Infect. Immun. 67:3267-3275 (1999)), an exclusion that led researchers away from the significance of CRP binding to respiratory pathogens. Although ChoP expression on bacteria has been found to confer resistance to antimicrobial peptides on the mucosa of the upper respirator tract, such as LL-36 and LL-37/hCAP18, that target structural differences in membranes between host and microbial cells (Lysenko et al., Infect. Immun. 68:1664-1671 (2000)), only the source and regulation of serum CRP were extensively studied.
The inventors were the first to examine the expression of CPR in the human respiratory tract, particularly in the heavily colonized upper respiratory tract where the organisms bearing the ChoP target reside (Gould & Weiser, Infect. Immun. 69(3):1747-1754 (2001)). Using a monoclonal antibody to CRP Gould and Weiser demonstrated that CRP is present in secretions from inflamed (0.17 to 42 μg/ml) and non-inflamed (<0.05 to 0.88 μg/ml) human respiratory tracts in sufficient quantities for an antimicrobial effect. In addition, they reported that the CRP gene was expressed in epithelial cells of the human respiratory tract using in situ hybridization on nasal polyps and reverse transcriptase PCR of pharyngeal cells in culture. However, there is phase variation in the presence or amount of ChoP on the bacterial cell surface in many species expressing this moiety. A direct antimicrobial or bactericidal effect of CRP has been demonstrated only for H. influenzae phase variants expressing ChoP in vitro, where concentrations of the protein as low as 20 ng/ml bind to ChoP and mediate a complement-dependent bactericidal effect (Weiser et al., J. Exp. Med., 1998, supra). Nevertheless, this finding only examined CRP in serum, offering no suggestion of direct antimicrobial binding to the pathogens in the respiratory tract.
Current diagnostic tests for respiratory tract infections are slow and often incorrect. The microbiological laboratory evaluation of an adult patient with a presumed respiratory tract infection may include a sputum gram stain, sputum culture and blood culture. Sputum cultures require over 24 hours before the infective organism(s) can be identified. Even then, however, the findings often reflect the patient's diverse oro-pharyngeal colonizing flora, which often may include known pathogens, rather than identify the organism causing the infection. Therefore, because of the high rate of error and lack of specificity, practitioners typically do not bother with sputum gram stains and cultures.
Blood tests are useful to determine microorganisms, if correct, but resolution also requires more than 24 hours of cell culture, and even then may be inaccurate. A blood culture when positive for a microorganism known to cause human respiratory tract infection is very helpful; however, blood cultures are infrequently positive even in the face of impressive radiographic evidence of pneumonia. Moreover, the time needed for cultures to mature in the laboratory can significantly delay establishing a microbiological diagnosis.
Consequently, physicians and healthcare practitioners frequently must rely upon laboratory results based upon current techniques that provide slow, and at best vague and often misleading information. This dilemma leaves the health care practitioner with the need to prognosticate the identity of the microorganism causing the respiratory infection in a patient, based empirically upon what infections are typical in a patient of the same age group with similar co-morbidities in light of the season in which the illness is occurring. The usual clinical scenario, therefore, is one in which the causative microorganism is never identified, and instead of delaying treatment, the patient is simply routinely treated with broad-spectrum antibiotics in the hope of achieving a clinical cure (Kim & Weiser, in Respiratory Infections, supra). This empiric usage of antibiotics has led to a serious antimicrobial resistance problem, in both the hospital and outpatient communities. This has been dramatically seen with Streptococcus pneumoniae, the most common respiratory tract pathogen in humans, which is increasingly selectively resistant to antibiotic treatment.
Accordingly, until the present invention there has remained a recognized need for a diagnostic test that will quickly and accurately determine the etiology of a respiratory tract infection in a patient to permit specific clinical treatment, as opposed to the frequent use of broad range antibiotics, which over time can induce serious antimicrobial resistance problems in the patient.